Method of removing non-specific binding signals using microparticle assay

Methods and kits for accurately detecting one or more analytes in a sample by removing non-specific binding signals utilizing capture and control microparticles. The capture microparticles can specifically bind to the analyte while the control microparticles do not specifically bind to the analyte but to the background molecules. Both capture and control microparticles are added to the sample under suitable conditions to allow binding between analytes and the microparticles. Detection agent is then added to bind to analytes and other substances captured by the microparticles. The microparticles are then run through a cytometry-based detection method, where detection signals from the capture and the control microparticles are distinguished. The differences between the detection signals from the capture and the control microparticles are obtained, which are then used to determine the presence and/or amounts of the analytes based on a previously determined relationship between such differences and known amount of the analyte.